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Gandasi, N. R., Yin, P., Riz, M., Chibalina, M. V., Cortese, G., Lund, P.-E., . . . Barg, S. (2017). Ca2+ channel clustering with insulin-containing granules is disturbed in type 2 diabetes. Journal of Clinical Investigation, 127(6), 2353-2364.
Open this publication in new window or tab >>Ca2+ channel clustering with insulin-containing granules is disturbed in type 2 diabetes
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2017 (English)In: Journal of Clinical Investigation, ISSN 0021-9738, E-ISSN 1558-8238, Vol. 127, no 6, 2353-2364 p.Article in journal (Refereed) Published
Abstract [en]

Loss of first-phase insulin secretion is an early sign of developing type 2 diabetes (T2D). Ca2+ entry through voltage-gated L-type Ca2+ channels triggers exocytosis of insulin-containing granules in pancreatic β cells and is required for the postprandial spike in insulin secretion. Using high-resolution microscopy, we have identified a subset of docked insulin granules in human β cells and rat-derived clonal insulin 1 (INS1) cells for which localized Ca2+ influx triggers exocytosis with high probability and minimal latency. This immediately releasable pool (IRP) of granules, identified both structurally and functionally, was absent in β cells from human T2D donors and in INS1 cells cultured in fatty acids that mimic the diabetic state. Upon arrival at the plasma membrane, IRP granules slowly associated with 15 to 20 L-type channels. We determined that recruitment depended on a direct interaction with the synaptic protein Munc13, because expression of the II-III loop of the channel, the C2 domain of Munc13-1, or of Munc13-1 with a mutated C2 domain all disrupted L-type channel clustering at granules and ablated fast exocytosis. Thus, rapid insulin secretion requires Munc13-mediated recruitment of L-type Ca2+ channels in close proximity to insulin granules. Loss of this organization underlies disturbed insulin secretion kinetics in T2D.

National Category
Cell and Molecular Biology
Research subject
Molecular Cellbiology
Identifiers
urn:nbn:se:uu:diva-321935 (URN)10.1172/JCI88491 (DOI)000402620800029 ()28481223 (PubMedID)
Funder
Swedish Research CouncilSwedish Diabetes AssociationThe Swedish Brain FoundationSwedish Child Diabetes FoundationEXODIAB - Excellence of Diabetes Research in SwedenNovo Nordisk
Available from: 2017-05-12 Created: 2017-05-12 Last updated: 2017-09-08Bibliographically approved
Marshall, M., Lund, P.-E. & Barg, S. (2017). Molecular Mechanisms of V-SNARE Function in Secretory Granule Exocytosis. Paper presented at 58th Annual Meeting of the Biophysical-Society, FEB 15-19, 2014, San Francisco, CA. Biophysical Journal, 112(3), 395A-395A.
Open this publication in new window or tab >>Molecular Mechanisms of V-SNARE Function in Secretory Granule Exocytosis
2017 (English)In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 112, no 3, 395A-395A p.Article in journal, Meeting abstract (Other academic) Published
Place, publisher, year, edition, pages
CELL PRESS, 2017
National Category
Biophysics
Identifiers
urn:nbn:se:uu:diva-332760 (URN)000402375600962 ()
Conference
58th Annual Meeting of the Biophysical-Society, FEB 15-19, 2014, San Francisco, CA
Available from: 2017-11-06 Created: 2017-11-06 Last updated: 2017-11-06
Barg, S. & Gucek, A. (2016). How Kiss-and-Run Can Make Us Sick: SOX4 Puts a Break on the Pore. Diabetes, 65(7), 1791-1793.
Open this publication in new window or tab >>How Kiss-and-Run Can Make Us Sick: SOX4 Puts a Break on the Pore
2016 (English)In: Diabetes, ISSN 0012-1797, E-ISSN 1939-327X, Vol. 65, no 7, 1791-1793 p.Article in journal, Editorial material (Other academic) Published
National Category
Endocrinology and Diabetes
Identifiers
urn:nbn:se:uu:diva-300622 (URN)10.2337/dbi16-0019 (DOI)000378463000005 ()27329955 (PubMedID)
Available from: 2016-08-10 Created: 2016-08-10 Last updated: 2017-11-28Bibliographically approved
Alenkvist, I., Gandasi, N. R., Barg, S. & Tengholm, A. (2015). Activation-dependent translocation of Epac2 to granule docking sites at the beta cell plasma membrane. Paper presented at 51st Annual Meeting of the European-Association-for-the-Study-of-Diabetes (EASD), SEP 14-18, 2015, Stockholm, SWEDEN. Diabetologia, 58(Suppl. 1), S210-S210.
Open this publication in new window or tab >>Activation-dependent translocation of Epac2 to granule docking sites at the beta cell plasma membrane
2015 (English)In: Diabetologia, ISSN 0012-186X, E-ISSN 1432-0428, Vol. 58, no Suppl. 1, S210-S210 p.Article in journal, Meeting abstract (Other academic) Published
National Category
Endocrinology and Diabetes
Identifiers
urn:nbn:se:uu:diva-264887 (URN)000359820901111 ()
Conference
51st Annual Meeting of the European-Association-for-the-Study-of-Diabetes (EASD), SEP 14-18, 2015, Stockholm, SWEDEN
Note

Meeting Abstract: 421

Available from: 2015-11-05 Created: 2015-10-19 Last updated: 2017-12-01Bibliographically approved
Yin, P., Gandasi, N. R., Riz, M., Cortese, G., Chibalina, M., Rorsman, P., . . . Barg, S. (2015). Clustering of L-type CA(2+)-channels promotes exocytosis of individual secretory granules. Paper presented at 51st Annual Meeting of the European-Association-for-the-Study-of-Diabetes (EASD), SEP 14-18, 2015, Stockholm, SWEDEN. Diabetologia, 58(Suppl. 1), S128-S128.
Open this publication in new window or tab >>Clustering of L-type CA(2+)-channels promotes exocytosis of individual secretory granules
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2015 (English)In: Diabetologia, ISSN 0012-186X, E-ISSN 1432-0428, Vol. 58, no Suppl. 1, S128-S128 p.Article in journal, Meeting abstract (Other academic) Published
National Category
Endocrinology and Diabetes
Identifiers
urn:nbn:se:uu:diva-264915 (URN)000359820900255 ()
Conference
51st Annual Meeting of the European-Association-for-the-Study-of-Diabetes (EASD), SEP 14-18, 2015, Stockholm, SWEDEN
Note

Meeting Abstract: 254

Available from: 2015-11-04 Created: 2015-10-19 Last updated: 2017-12-01Bibliographically approved
Kay, E. I. & Barg, S. (2015). G-protein coupled receptor dynamics in insulin secreting cells. Paper presented at 51st Annual Meeting of the European-Association-for-the-Study-of-Diabetes (EASD), SEP 14-18, 2015, Stockholm, SWEDEN. Diabetologia, 58(Suppl. 1), S212-S212.
Open this publication in new window or tab >>G-protein coupled receptor dynamics in insulin secreting cells
2015 (English)In: Diabetologia, ISSN 0012-186X, E-ISSN 1432-0428, Vol. 58, no Suppl. 1, S212-S212 p.Article in journal, Meeting abstract (Other academic) Published
National Category
Endocrinology and Diabetes
Identifiers
urn:nbn:se:uu:diva-264903 (URN)000359820901116 ()
Conference
51st Annual Meeting of the European-Association-for-the-Study-of-Diabetes (EASD), SEP 14-18, 2015, Stockholm, SWEDEN
Note

Meeting Abstract: 426

Available from: 2015-11-05 Created: 2015-10-19 Last updated: 2017-12-01Bibliographically approved
Salunkhe, V. A., Ofori, J., Gandasi, N. R., Salo, S. A., Hansson, S., Andersson, M. E., . . . Eliasson, L. (2015). MiR-335 regulates exocytotic proteins and affects glucose-stimulated insulin secretion through decreased Ca2+-dependent exocytosis in beta cells. Paper presented at 51st Annual Meeting of the European-Association-for-the-Study-of-Diabetes (EASD), SEP 14-18, 2015, Stockholm, SWEDEN. Diabetologia, 58(Suppl. 1), S128-S128.
Open this publication in new window or tab >>MiR-335 regulates exocytotic proteins and affects glucose-stimulated insulin secretion through decreased Ca2+-dependent exocytosis in beta cells
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2015 (English)In: Diabetologia, ISSN 0012-186X, E-ISSN 1432-0428, Vol. 58, no Suppl. 1, S128-S128 p.Article in journal, Meeting abstract (Other academic) Published
National Category
Endocrinology and Diabetes
Identifiers
urn:nbn:se:uu:diva-264912 (URN)000359820900256 ()
Conference
51st Annual Meeting of the European-Association-for-the-Study-of-Diabetes (EASD), SEP 14-18, 2015, Stockholm, SWEDEN
Note

Meeting Abstract: 255

Available from: 2015-11-04 Created: 2015-10-19 Last updated: 2017-12-01Bibliographically approved
Gandasi, N. R. & Barg, S. (2015). Quantitative Imaging of the Exocytosis Machinery Assembly. Biophysical Journal, 108(2), 102A-102A.
Open this publication in new window or tab >>Quantitative Imaging of the Exocytosis Machinery Assembly
2015 (English)In: Biophysical Journal, ISSN 0006-3495, E-ISSN 1542-0086, Vol. 108, no 2, 102A-102A p.Article in journal, Meeting abstract (Other academic) Published
National Category
Biophysics
Identifiers
urn:nbn:se:uu:diva-274842 (URN)000359471700514 ()
Available from: 2016-01-26 Created: 2016-01-26 Last updated: 2017-11-30Bibliographically approved
Gandasi, N. R. & Barg, S. (2015). Rabs and RIM are differentially involved in docking and tethering of insulin granules. Paper presented at 51st Annual Meeting of the European-Association-for-the-Study-of-Diabetes (EASD), SEP 14-18, 2015, Stockholm, SWEDEN. Diabetologia, 58(Suppl. 1), S127-S127.
Open this publication in new window or tab >>Rabs and RIM are differentially involved in docking and tethering of insulin granules
2015 (English)In: Diabetologia, ISSN 0012-186X, E-ISSN 1432-0428, Vol. 58, no Suppl. 1, S127-S127 p.Article in journal, Meeting abstract (Other academic) Published
National Category
Endocrinology and Diabetes
Identifiers
urn:nbn:se:uu:diva-264897 (URN)000359820900254 ()
Conference
51st Annual Meeting of the European-Association-for-the-Study-of-Diabetes (EASD), SEP 14-18, 2015, Stockholm, SWEDEN
Note

Meeting Abstract: 253

Available from: 2015-11-05 Created: 2015-10-19 Last updated: 2017-12-01Bibliographically approved
Gandasi, N. R., Vesto, K., Helou, M., Yin, P., Saras, J. & Barg, S. (2015). Survey of Red Fluorescence Proteins as Markers for Secretory Granule Exocytosis. PLoS ONE, 10(6), Article ID e0127801.
Open this publication in new window or tab >>Survey of Red Fluorescence Proteins as Markers for Secretory Granule Exocytosis
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2015 (English)In: PLoS ONE, ISSN 1932-6203, E-ISSN 1932-6203, Vol. 10, no 6, e0127801Article in journal (Refereed) Published
Abstract [en]

Fluorescent proteins (FPs) have proven to be valuable tools for high-resolution imaging studies of vesicle transport processes, including exo- and endocytosis. Since the pH of the vesicle lumen changes between acidic and neutral during these events, pH-sensitive FPs with near neutral pKa, such as pHluorin, are particularly useful. FPs with pKa>6 are readily available in the green spectrum, while red-emitting pH-sensitive FPs are rare and often not well characterized as reporters of exo- or endocytosis. Here we tested a panel of ten orange/red and two green FPs in fusions with neuropeptide Y (NPY) for use as secreted vesicle marker and reporter of dense core granule exocytosis and release. We report relative brightness, bleaching rate, targeting accuracy, sensitivity to vesicle pH, and their performance in detecting exocytosis in live cells. Tandem dimer (td)-mOrange2 was identified as well-targeted, bright, slowly bleaching and pH-sensitive FP that performed similar to EGFP. Single exocytosis events were readily observed, which allowed measurements of fusion pore lifetime and the dynamics of the exocytosis protein syntaxin at the release site during membrane fusion and cargo release.

National Category
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
Identifiers
urn:nbn:se:uu:diva-258769 (URN)10.1371/journal.pone.0127801 (DOI)000356835000012 ()26091288 (PubMedID)
Funder
Swedish Research CouncilSwedish Diabetes Association
Available from: 2015-07-20 Created: 2015-07-20 Last updated: 2017-12-04Bibliographically approved
Organisations
Identifiers
ORCID iD: ORCID iD iconorcid.org/0000-0003-4661-5724

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